The Sequencing facility provides VUMC and VU investigators with automated DNA sequencing technology on a fee for service basis. The sequencing is performed on an ABI 3730 DNA analyzer that uses capillary technology to achieve high- throughput sequencing.
The facility offers three sequencing services: Ready-to-Run Service, Full Service and Customised service.
The client performs the sequencing reactions and brings in the purified samples for running on the capillary sequencer. Samples can be delivered in microcentrifuges tubes or in a PCR plate in case of large number of samples (we provide you directly with an optical bar coded plate).
In the ready-to-run service the facility does not perform additional processing of client samples in any way. The facility simply pipettes the samples in an optical plate according to the sample-scheme provided by the client and then the plate is loaded into the capillary sequencer. To verify that all equipment is functioning correctly regularly validation checks are run on all instruments with control samples. Therefore, if the run is successful for other samples on the plate and a single clients samples fail, the facility will have to decline any responsibility for the failure but personnel is available for troubleshooting.
The facility carries out the sequencing reactions on sample for the client.
Samples are accepted in two formats: plasmids and PCR products
Please note that quality of sequencing is directly proportional to the quality of the DNA.
Therefore to insure the success of your sequencing reactions the client has to make sure that the DNA is of good quality and the concentration is accurately measured. The client needs to quantitate plasmids and PCR products on an agarose gel next to a DNA standard (e.g. low mass and /or high mass standard from Invitrogen for example). If requested the facility will run the agarose gel for the client. For plasmids and PCR products the facility requests primers in 0,5 or 1,5 ml eppendorf tubes clearly labelled with the name of the primers at the concentration of 10 pmol/ul.
The facility also performs complete sequencing projects according to the client needs including PCR, PCR purification and sequencing reactions; please enquire about the possibilities with the responsible person in the facility. Because of the unique characteristics of each project, this service is a customized process. The facility will discuss with the inquiring investigator the proper design and goals of the project. During this meeting the most appropriate technology for the specific project and an estimated cost is determined.
Ready-to-run reactions: 0,70 euro reaction for Clinical Genetics investigators; 1,25 euro for other investigators from VUmc and VU departments on the basis of 48 or 96 wells formats.
Please remember; 1 sequencing reaction is one template with one primer!!
For ready to run sequences please inform the facility by email (firstname.lastname@example.org) at least one day in advance clearly specifying how many samples you want to run. Once you receive a confirmation email drop your samples at the facility directly in the fridge in room A-327 in the box marked as: sequencing for guest and email us the correct ABI plate record template. Click here to open: The Word document README and the Excel files: plate record seq-384-v3-IUB ; plate record seq-96-PCR-v3 ; plate record seq-96-clones-v3 ; plate record GS-96-G5-LIZ ; plate record GS-384-G5-LIZ )
Turn-around time for the ready-to-run reactions in 48 or 96 wells formats is 1-2 working days. If you have less than 48 samples you might decide to wait until we reach a full run (48 samples) or run your samples anyway and pay the full charge (48 samples). Turn-around time for the full service will depend on the size of the project and will be indicated per project.
Results will be available as ABI computer files and they can be retrieved either directly from the facility or from the VUmc intranet connection if you warrant us access.